Tagging quantitative loci controlling pathogenicity in Magnaporthe grisea by insertional mutagenesis

2002 ◽  
Vol 61 (2) ◽  
pp. 77-88 ◽  
Author(s):  
D. Fujimoto ◽  
Y. Shi ◽  
D. Christian ◽  
J.B. Mantanguihan ◽  
H. Leung
Keyword(s):  
Insertional Mutagenesis ◽  
Magnaporthe Grisea

scholarly journals Insertion of the LINE Retrotransposon MGL Causes a Conidiophore Pattern Mutation in Magnaporthe grisea

2000 ◽  
Vol 13 (8) ◽  
pp. 892-894 ◽  
Author(s):  
Marie Nishimura ◽  
Nagao Hayashi ◽  
Nam-Soo Jwa ◽  
Gee W. Lau ◽  
John E. Hamer ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Aspergillus Nidulans ◽  
Insertional Mutagenesis ◽  
Magnaporthe Grisea ◽  
Morphological Mutants

We obtained three Magnaporthe grisea morphological mutants that had the LINE transposon MGL inserted into the ACR1 locus. Sequence analysis revealed that ACR1 is homologous to medA, a developmental regulator of Aspergillus nidulans conidiation. These results demonstrated that MGL elements could transpose and cause insertional mutagenesis in M. grisea.

scholarly journals Transposon impala, a Novel Tool for Gene Tagging in the Rice Blast Fungus Magnaporthe grisea

2001 ◽  
Vol 14 (3) ◽  
pp. 308-315 ◽  
Author(s):  
François Villalba ◽  
Marc-Henri Lebrun ◽  
Aurélie Hua-Van ◽  
Marie-Josée Daboussi ◽  
Marie-Claire Grosjean-Cournoyer
Keyword(s):  
Nitrate Reductase ◽  
Fusarium Oxysporum ◽  
Rice Blast ◽  
Insertional Mutagenesis ◽  
Magnaporthe Grisea ◽  
Rice Blast Fungus ◽  
Single Copy ◽  
Wild Type ◽  
Type Locus ◽  
Blast Fungus

impala, a Tc1-mariner transposable element from Fusarium oxysporum, was introduced into the rice blast fungus Magnaporthe grisea to develop transposon-based insertional mutagenesis. A construct (pNIL160) containing an autonomous impala copy inserted in the promoter of niaD encoding Aspergillus nidulans nitrate reductase was introduced by transformation into a M. grisea nitrate reductase-deficient mutant. impala excision was monitored by restoration of prototrophy for nitrate. Southern analysis of niaD+ revertants revealed that impala was able to excise and reinsert at new loci in M. grisea. As observed for its host Fusarium oxysporum, impala inserted at a TA site left a typical excision footprint of 5 bp. We have shown that a defective impala copy was inactive in M. grisea, yet it can be activated by a functional impala transposase. A transformant carrying a single copy of pNIL160 was used to generate a collection of 350 revertants. Mutants either altered for their mycelial growth (Rev2) or nonpathogenic (Rev77) were obtained. Complementation of Rev77 with a 3-kb genomic fragment from a wild-type locus was successful, demonstrating the tagging of a pathogenicity gene by impala. This gene, called ORP1, is essential for penetration of host leaves by M. grisea and has no sequence homology to known genes.

scholarly journals Identification of Pathogenicity Mutants of the Rice Blast Fungus Magnaporthe grisea by Insertional Mutagenesis

1999 ◽  
Vol 12 (2) ◽  
pp. 129-142 ◽  
Author(s):  
Pascale V. Balhadère ◽  
Andrew J. Foster ◽  
Nicholas J. Talbot
Keyword(s):  
Insertional Mutagenesis ◽  
Magnaporthe Grisea ◽  
Single Gene ◽  
Phenotypic Analysis ◽  
Dependent Protein Kinase ◽  
Dna Integration ◽  
Gene Segregation ◽  
Similar Phenotype ◽  
Dependent Protein ◽  
Remi Mutagenesis

Restriction enzyme-mediated DNA integration (REMI) mutagenesis was used to identify mutants of Magnaporthe grisea impaired in pathogenicity. Three REMI protocols were evaluated and the frequency of REMIs determined. An REMI library of 3,527 M. grisea transformants was generated in three genetic backgrounds, and 1,150 transformants were screened for defects in pathogenicity with a barley cut leaf assay. Five mutants were identified and characterized. Two mutants (2029 and 2050) were impaired in appressorium function. Two other mutants, 125 and 130, were altered in conidial morphology, conidiogenesis, and appressorium function. Mutant 130 was also a methionine auxotroph and methionine auxotrophy co-segregated with the reduction in pathogenicity. An additional mutant, 80, showed reduced pathogenicity on blast-susceptible rice cultivars but was fully pathogenic on barley. The reduction of pathogenicity in mutant 80 was associated with a delay in conidial germination and appressorium development. Genetic analysis suggested single-gene segregation for each mutant, but only two of the mutations co-segregated with the hygromycin resistance marker. The genetic loci in mutants 2029, 2050, 125, 130, and 80 were termed PDE1, PDE2, IGD1, MET1, and GDE1, respectively. pde1 and pde2 were non-allelic to cpkA, a mutation in the catalytic subunit of cyclic AMP (cAMP)-dependent protein kinase A with a very similar phenotype. The results indicate the utility of REMI for studying fungal pathogenicity, but also highlight the requirement for rigorous genetic and phenotypic analysis.

scholarly journals Magnaporthe grisea Pathogenicity Genes Obtained Through Insertional Mutagenesis

1998 ◽  
Vol 11 (5) ◽  
pp. 404-412 ◽  
Author(s):  
James A. Sweigard ◽  
Anne M. Carroll ◽  
Leonard Farrall ◽  
Forrest G. Chumley ◽  
Barbara Valent
Keyword(s):  
Restriction Enzyme ◽  
Rice Blast ◽  
Primary Infection ◽  
Insertional Mutagenesis ◽  
Magnaporthe Grisea ◽  
Mutational Analysis ◽  
Rice Blast Fungus ◽  
Hygromycin Resistance ◽  
Resistance Marker ◽  
Pathogenicity Genes

We have initiated a mutational analysis of pathogenicity in the rice blast fungus, Magnaporthe grisea, in which hygromycin-resistant transformants, most generated by restriction enzyme-mediated integration (REMI), were screened for the ability to infect plants. A rapid primary infection assay facilitated screening of 5,538 transformants. Twenty-seven mutants were obtained that showed a reproducible pathogenicity defect, and 18 of these contained mutations that cosegregated with the hygromycin resistance marker. Analysis of eight mutants has resulted in the cloning of seven PTH genes that play a role in pathogenicity on barley, weeping lovegrass, and rice. Two independent mutants identified the same gene, PTH2, suggesting nonrandom insertion of the transforming DNA. These first 7 cloned PTH genes are described.

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